Sulfonation occurs as a common enzymatic modification of endogenous substances including proteins, carbohydrates, catecholamines, and estrogenic steroids as well as xenobiotic chemicals [1]. Sulfonation refers to the transfer of the sulfonate group (SO3−1) from 3′-phosphoadenosine-5′-phosphosulfate (PAPS), the only known sulfonate donor [2]. Sulfonation can occur through several types of linkages, esters (O-sulfonation), amides (N-sulfonation) and thioesters (S-sulfonation) [3], of which O-sulfonation is the most prominent. The transfer of SO3−1 to a hydroxyl acceptor (O-sulfonation) generates an ester sulfate, and this reaction has commonly been referred to as sulfation rather than the more accurate O-sulfonation.
The majority of cellular sulfonation is O-sulfonation and occurs primarily on steroids, catecholamines and thyroid hormones [1]. The sulfonation of these molecules is catalyzed by the soluble cytosolic sulfotransferases and appears to alter their bioactivity. For example, estrogen, testosterone and thyroid hormones (T3 and T4) can interact with their respective receptors to regulate transcription whereas their sulfate-containing moieties cannot. Furthermore, the half-life of these compounds in blood is significantly shorter than that of their conjugated counterparts suggesting that sulfonation maintains these compounds in an inactive state ready for rapid deployment by the removal of the sulfonyl group.
While the cytosolic sulfotransferases conjugate cell-permeable or intracellular compounds, the membrane-bound Golgi-associated sulfotransferases are primarily responsible for sulfonation of extracellular proteins via a co- or post-translational mechanism. The membrane-bound sulfotransferases are responsible for the sulfonation of various glycosaminoglycans such as heparin and heparan sulfate. Additionally, such enzymes catalyze the direct sulfonation of proteins on the 4-O-position of tyrosine residues [4]. It is one of the last modifications to occur during protein transiting the trans-Golgi and thus has been found almost exclusively on secreted and plasma membrane proteins of all metazoan species examined. In addition, there is a large body of evidence that this modification is present usually at the interface of interacting proteins and hence, is known to modulate extracellular protein-protein interactions. In humans, protein tyrosine sulfonation has been implicated in proteins of the vasculature and hemostasis. Examples include the mediation of inflammatory leukocyte adhesion, chemokine receptors and modulation of the blood coagulation cascade [5]. Significantly, only tyrosine residues have been described as sites for O-sulfonation within proteins, and O-sulfonation of proteins has not previously been shown to occur within the cytosol. Several tyrosyl protein sulfotransferases [6,7] and arylsulfatases [8] present in the trans-Golgi have been described, but unlike tyrosine phosphorylation/dephosphorylation [9], there is no evidence of dynamic regulation of tyrosine sulfonation [4,5]. Until now, only widespread modification of tyrosine has been observed [10, 11].
We have discovered the occurrence of sulfonation as a posttranslational modification of serine and threonine residues, and that this sulfonation is involved in the modulation of protein-protein interactions, and in particular, has regulatory functions in receptor tyrosine kinase signaling as discussed further below. We have exploited this finding to develop methods of detecting and modulating this serine and threonine sulfonation.